Nagaoka M H, Maitani T
National Institute of Health Sciences, Kamiyoga 1-18-1, Setagaya, 158-8501, Tokyo, Japan.
Biochim Biophys Acta. 2000 Oct 18;1523(2-3):182-8. doi: 10.1016/s0304-4165(00)00120-3.
The binding of iron (Fe) to human serum transferrin (Tf) was analyzed with an HPLC system equipped with an anion exchange column and directly connected with a high-resolution inductively coupled plasma mass spectrometer for metal detection. The (56)Fe level in the eluate was monitored at resolution m/Deltam=3000. Two monoferric Tfs were assigned based on the results of urea-PAGE and desferrioxamine experiments. When Fe was added as Fe-citrate stepwise to an apo-Tf solution in the presence of bicarbonate, the N-lobe site was the preferential Fe-binding site, while the C-lobe site was preferred in the absence of bicarbonate. In both cases, the Fe-peak areas of the preferential site and Fe(2)-Tf increased up to an Fe/Tf molar ratio of 1, and then the peak area of the monoferric Tf decreased while the peak area of Fe(2)-Tf increased. When the Fe/Tf molar ratio was below 1, the amount of Fe bound to the lobe with a weaker affinity was higher in Fe(2)-Tf than in the monoferric Tf in each case. Namely, Fe(2)-Tf was the preferential binding state of Fe to human serum Tf. The preference is reasonable for transferring Fe ions effectively to Tf-receptors.
采用配备阴离子交换柱并直接连接高分辨率电感耦合等离子体质谱仪进行金属检测的高效液相色谱系统，分析铁（Fe）与人血清转铁蛋白（Tf）的结合情况。在分辨率m/Δm = 3000下监测洗脱液中的（56）Fe水平。根据尿素聚丙烯酰胺凝胶电泳和去铁胺实验结果确定了两种单铁转铁蛋白。当在碳酸氢盐存在下将柠檬酸铁逐步添加到脱铁转铁蛋白溶液中时，N叶位点是优先的铁结合位点，而在没有碳酸氢盐的情况下C叶位点是优先的。在这两种情况下，优先位点的铁峰面积和Fe(2)-Tf的铁峰面积在铁/转铁蛋白摩尔比达到1之前均增加，然后单铁转铁蛋白的峰面积减小而Fe(2)-Tf的峰面积增加。当铁/转铁蛋白摩尔比低于1时，在每种情况下，Fe(2)-Tf中与亲和力较弱的叶结合的铁量均高于单铁转铁蛋白。即，Fe(2)-Tf是铁与人血清转铁蛋白的优先结合状态。这种偏好对于将铁离子有效地转移到转铁蛋白受体来说是合理的。
